Impact of Nosema ceranae invasion on sucrose solution consumption, midgut epithelial cell structure, and lifespan of Apis cerana cerana workers

Nosema ceranae is an intracellular fungal parasite for honeybees, leading to chronic disease named bee nosemosis with worldwide distribution. Asian honeybee (Apis cerana) is the original host for N. ceranae, but the impact of N. ceranae infection on A. cerana physiology is largely unknown. In this current work, workers of Apis cerana cerana, a subspecies of Asian honeybee, were artificially inoculated with N. ceranae spores and reared under lab conditions, followed by detection of fungal spore load as well as host sucrose solution consumption, midgut epithelial cell structure, and lifespan. The result of spore counting suggested that the spore load in the host midgut decreased significantly during 1 dpi-2 dpi, whereas that displayed an elevated trend among 2 dpi-13 dpi. The sucrose solution consumption of workers in N. ceranae-inoculated groups among 1 dpi-20 dpi was always higher than that of workers in un-inoculated groups; additionally, the difference of sucrose solution consumption between these two groups at 4 dpi, 5 dpi, and 13 dpi was of significance. Based on microscopic observation of paraffin sections, darkly stained parasites were clearly detected in the midgut epithelial cells of N. ceranae-inoculated workers at 7 dpi-10 dpi, whereas no parasite was observed in those of un-inoculated workers. In addition, the boundaries of un-inoculated host epithelial cells were intact and the darkly stained nucleus were clear, while the boundaries of midgut epithelial cells of N. ceranae-inoculated workers were blurred, the nucleus were almost disappeared, and the nucleic acid substances were diffused. Moreover, the survival rates of workers in both N. ceranae-inoculated groups and un-inoculated groups at 1 dpi-5 dpi were pretty high and then started to decrease at 5 dpi; the survival rate of workers in N. ceranae-inoculated groups was always lower than that in un-inoculated groups, with significant difference between these two groups during 11 dpi-20 dpi. These results together indicate that the quantity of fungal spores continuously elevated with the microsporidian multiplication, causing energetic stress for workers and host cell structure damage, which further negatively affected the host lifespan. Our findings offer a solid basis not only for exploring the molecular mechanism underlying N. ceranae infection but also for investigating the interaction between N. ceranae and eastern honeybee.

Genome Sequence of the Asian Honeybee in Pakistan Sheds Light on Its Phylogenetic Relationship with Other Honeybees

In Pakistan, Apis cerana, the Asian honeybee, has been used for honey production and pollination services. However, its genomic makeup and phylogenetic relationship with those in other countries are still unknown. We collected A. cerana samples from the main cerana-keeping region in Pakistan and performed whole genome sequencing. A total of 28 Gb of Illumina shotgun reads were generated, which were used to assemble the genome. The obtained genome assembly had a total length of 214 Mb, with a GC content of 32.77%. The assembly had a scaffold N50 of 2.85 Mb and a BUSCO completeness score of 99%, suggesting a remarkably complete genome sequence for A. cerana in Pakistan. A MAKER pipeline was employed to annotate the genome sequence, and a total of 11,864 protein-coding genes were identified. Of them, 6750 genes were assigned at least one GO term, and 8813 genes were annotated with at least one protein domain. Genome-scale phylogeny analysis indicated an unexpectedly close relationship between A. cerana in Pakistan and those in China, suggesting a potential human introduction of the species between the two countries. Our results will facilitate the genetic improvement and conservation of A. cerana in Pakistan.

Composition and diversity of bacteria from giant Asian honeybee Apis dorsata gut

Niode NJ, Adji A, Rimbing J, Tulung M, Tallei TE. 2021. Composition and diversity of bacteria from giant Asian honeybee Apis dorsata gut. Biodiversitas 22: 906-912. As a social insect, honeybee possesses a unique gut bacteria community. Therefore, we need to understand the composition and diversity of bacteria in Apis dorsata gut, the largest honeybee species that live in the forest, especially in Southeast Asia. The present study aimed to investigate the gut bacteria of A. dorsata using a metabarcoding approach. The DNA barcode region used in this approach was the V3-V4 region of 16S rRNA. Honeybees were caught in a forest area located in Tareran, South Minahasa, North Sulawesi, Indonesia. There were 11 phyla identified from the gut of A. dorsata. The most abundant phyla were Firmicutes (95.8%), Proteobacteria (3.7%), and Actinobacteria (0.4%). The class Bacilli was responsible for 94.5% of the total bacterial population, the dominant family was Bacillaceae (87.2%), and the dominant genus was Bacillus (87%). Simpson (1-D) 0.24 and Shannon diversity index 0.98 indicated that the diversity of the genera was low. A nonsignificant number of species belong to the lactic acid bacteria was also detected, which may have certain benefits that need to be investigated further.

Seasonal prevalence of Asian honeybee ectoparasitic mite Varroa destructor Anderson and Trueman, 2000 in Madanpokhara Apiaries, Palpa, Nepal

The present study highlights the prevalence and seasonal variation of Varroa destructor Anderson and Trueman, 2000 in Apis cerana colonies at two apiaries (site-I and site-II) in Madanpokhara, Palpa district, Nepal. Altogether 498 varroa mites were collected from February to October 2017. The highest and lowest number of varroa mites were observed in March and September, respectively. The mites were observed higher in brood cells (54.84% and 52.51%) and lower in adult bees (10.39% and 9.59%) in site-I and site-II, respectively. The inner hive and outer temperature in site-I and II were ranged from 30°C to 34°C and 24°C to 32.4°C, 30°C to 33.5°C and 25.1°C to 32.3°C, respectively. Similarly, inner and outer hive humidity ranged from 41% to 61% and 40% to 68%, 40% to 60% and 41% to 65% at site-I and II respectively, differed significantly and correlated positively with mite population. Concern for honeybee health and conservation along with Nepali apiculture are also discussed in this paper.

Gene reuse facilitates rapid radiation and independent adaptation to diverse habitats in the Asian honeybee

Animals with recent shared ancestry frequently adapt in parallel to new but similar habitats, a process often underlined by repeated selection of the same genes. Yet, in contrast, few examples have demonstrated the significance of gene reuse in colonization of multiple disparate habitats. By analyzing 343 genomes of the widespread Asian honeybee, Apis cerana, we showed that multiple peripheral subspecies radiated from a central ancestral population and adapted independently to diverse habitats. We found strong evidence of gene reuse in the Leucokinin receptor (Lkr), which was repeatedly selected in almost all peripheral subspecies. Differential expression and RNA interference knockdown revealed the role of Lkr in influencing foraging labor division, suggesting that Lkr facilitates collective tendency for pollen/nectar collection as an adaptation to floral changes. Our results suggest that honeybees may accommodate diverse floral shifts during rapid radiation through fine-tuning individual foraging tendency, a seemingly complex process accomplished by gene reuse.

Quantitative Measurement of Melittin in Asian Honeybee Venom Using a New Method Including UPLC-QqTOF-MS

Asian honeybee venom is widely used in traditional oriental medicine. Melittin is the main component of Asian honeybee venom. In the present study, an ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QqTOF-MS) method was used for accurate qualitative and quantitative analyses of melittin in Asian honeybee venom. The results showed that the dynamic linear range of melittin was from 0.094 to 20 μg/mL, and the limit of quantification was 0.3125 μg/mL. The spiking recovery of melittin in honeybee venom ranged from 84.88% to 93.05%. Eighteen Asian honeybee venom samples in eighteen batches were collected from two different zones of China, and their melittin contents were measured. The contents of melittin in Asian honeybee venom samples was 33.9–46.23% of dry weight. This method proved a useful tool for the rapid evaluation of the authenticity and quality of Asian honeybee venom in terms of the melittin contents, and will contribute to a broader understanding of Asian honeybee venom.